A Bacillus subtilis gene encoding a protein similar to nucleotide sugar transferases influences cell shape and viability

Abstract

Bacillus subtilis gene ypfP, which is located at 196°on the genetic map, shows similarity to both the monogalactosyldiacylglycerol synthase gene of Cucumis sativus, which encodes a galactosyltransferase, and the murG genes of B. subtilis, Escherichia coli, Haemophilus influenzae, and Synechocystis sp. strain PCC6803, which encode N-acetylglucosaminyltransferases involved in peptidoglycan biosynthesis. Cells containing a null mutation of ypfP are shorter and rounder than wild-type cells during growth in Luria-Bertani medium and glucose minimal medium. In addition, the mutant cells preferentially undergo lysis when grown on solid Luria-Bertani medium.The chloroplast membranes of higher plants and eukaryotic algae contain two major galactolipids, monogalactodiacylglycerol (MGDG) and digalactosyldiacylglycerol. The final step in the synthesis of MGDG is catalyzed by MGDG synthase, a UDPgalactose:1,2-diacylglycerol 3-␤-D-galactosyltransferase, which transfers galactose from UDPgalactose to 1,2-diacylglycerol (9). A sequence of an MGDG synthase from cucumber (Cucumis sativus) is known and was shown to be similar to those of the MurG peptidoglycan biosynthesis enzymes of Escherichia coli and Bacillus subtilis (17). The rigid peptidoglycan layer of the bacterial cell wall is composed of long glycan chains that are cross-linked through peptide side chains. The glycan chains are alternating copolymers of N-acetylglucosamine (GlcNAc) and N-acetylmuramic acid (MurNAc) that are held together by ␤134 glycosidic linkages. The chains are formed by polymerization of disaccharide units that are assembled on a lipid carrier molecule at the inner surface of the cytoplasmic membrane (11,12,18). Synthesis of the disaccharide is catalyzed by MurG, a UDP-N-acetylglucosamine:Nacetylmuramyl-(pentapeptide) pyrophosphoryl-undecaprenol N-acetylglucosamine transferase, which transfers GlcNAc from the nucleotide sugar UDP-GlcNAc to undecaprenyl-pyrophosphoryl-MurNAc-pentapeptide to form undecaprenylpyrophosphoryl-MurNAc-(pentapeptide)GlcNAc (2, 14). Here we report that B. subtilis contains a gene, ypfP, that influences cell shape and viability, and whose inferred product resembles MGDG synthase of C. sativus and shows similarity to some conserved regions of MurG proteins.ypfP, which is 382 codons in length, was previously identified by the B. subtilis sequencing project and lies between metB and cspD at 196°on the chromosome (3). Our searches against available sequence databases have revealed that the inferred amino acid sequence of YpfP is most similar to that of MGDG synthase of C. sativus. An alignment of these proteins shows they exhibit 24% identity overall and 37% identity in four regions involving 183 YpfP residues (Fig. 1). In confirmation and extension of the results obtained by Shimojima et al. (17), we found that YpfP, like MGDG synthase, contains regions of great similarity to some conserved regions of the MurG proteins of B. subtilis (1,7,15), Haemophilus influenzae (5), E. coli (8,13,19), and Synechocystis sp. strain PCC6803 (10...

Abnormal morphology of Bacillus subtilis ugtP mutant cells lacking glucolipids

“…Previously, ugtP mutant cells have already been reported to be shorter and rounder than the wild type (Price et al, 1997). However, we observed a different morphology.…”

“…These are synthesized processively by UgtP, which transfers glucose to diacylglycerol from UDP-glucose (Jorasch et al, 1998). The phenotype of cells with ugtP (ypfP) disruption has been reported to be shorter and rounder for the first time by Price et al (1997). The ugtP mutant cells have also been reported to show strikingly abnormal cell morphology (Lazarevic et al, 2005).…”

Configuration of the sugar head of glycolipids in thylakoid membranes

“…A knockout mutant of this gene in Bacillus subtilis had a round phenotype (Price et al, 1997;Matsuoka et al, 2011). Detailed regulatory mechanisms of this phenotype have been reported (Seki et al, 2015;Matsuoka et al, 2016) and summarized (Matsuoka, 2018 Introduction of ypfP/ugtP of S. aureus (ugt106B1) into Synechococcus was also attempted, but no transformants were obtained (Hölzl et al, 2005b).…”

The <i>Bacillus subtilis</i> essential gene <i>dgkB</i> is dispensable in mutants with defective lipoteichoic acid synthesis

“…The genes without GFPfusion were designed to be expressed using the RBS on the vector. The allele ugtP::kan in which ugtP was replaced with a kan cassette was from strain KP261 (Price et al, 1997).…”